Publications (5)4.82 Total impact
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Article: A robust method to quantify low molecular weight contaminants in heparin: detection of tris(2-n-butoxyethyl) phosphate
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ABSTRACT: Recently, oversulfated chondroitin sulfate (OSCS) was identified in contaminated heparin preparations, which were linked to several adverse clinical events and deaths. Orthogonal analytical techniques, namely nuclear magnetic resonance (NMR) and capillary electrophoresis (CE), have since been applied by several authors for the evaluation of heparin purity and safety. NMR identification and quantification of residual solvents and non-volatile low molecular contaminants with USP acceptance levels of toxicity was achieved 40-fold faster than the traditional GC-headspace technique, which takes ~120 min against ~3 min to obtain a (1)H NMR spectrum with a signal/noise ratio of at least 1000/1. The procedure allowed detection of Class 1 residual solvents at 2 ppm and quantification was possible above 10 ppm. 2D NMR techniques (edited-HSQC (1)H/(13)C) permitted visualization of otherwise masked EDTA signals at 3.68/59.7 ppm and 3.34/53.5 ppm, which may be overlapping mononuclear heparin signals, or those of ethanol and methanol. Detailed NMR and ESI-MS/MS studies revealed a hitherto unknown contaminant, tris(2-n-butoxyethyl) phosphate (TBEP), which has potential health risks.The Analyst. 01/2011; 136(11):2330-8. -
Article: Some biomolecules and a partially O-acetylated exo-galactomannan containing β-Galf units from pathogenic Exophiala jeanselmei, having a pronounced immunogenic response.
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ABSTRACT: The pathogenic fungus Exophiala jeanselmei (Ej4) was grown in submerged MM medium, glucose being consumed after six days with maximum biomass and EPS production. Cells were extracted with CHCl3-MeOH (2:1, v/v) yielding a product containing 10% lipid, with high levels of unsaturated C(18:1) (43.6%) and C(18:2) (21.0%), 2D-TLC showed the presence of PE (17.7%), PS (11.6%), PC (35.8%), PI (1.2%) and lyso-phospholipids, LPE (10.7%), LPC (2.0%), PA (10.4%), cardiolipin (10.5%) and glucosyl-ceramide. Analysis of EPS-1 (120 kDa) showed a galactomanan, containing a main chain of Manp-(1→2) (24.2%), substituted by side chains containing terminal Galf (16.8%) and Manp (3.5%) and acetyl groups attached at O-6 of terminal Galf. An immune response against antigens was obtained using Balb/C mice. Anti-EPS-1 antibodies recognized purified fraction containing cellular walls very titer and higher than 1:20,000 for EPS. The studied biomolecules showed biotechnological potential and point to important perspectives in diagnosis of fungi and immunomodulatory products.International journal of biological macromolecules 11/2010; 48(1):177-82. · 2.37 Impact Factor -
Article: The opportunistic fungal pathogen Scedosporium prolificans: carbohydrate epitopes of its glycoproteins.
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ABSTRACT: Isolated from the mycelium of Scedosporium prolificans were complex glycoproteins (RMP-Sp), with three structurally related components (HPSEC). RMP-Sp contained 35% protein and 62% carbohydrate with Rha, Ara, Man, Gal, Glc, and GlcNH(2) in a 18:1:24:8:6:5 molar ratio. Methylation analysis showed mainly nonreducing end- of Galp (13%), nonreducing end- (9%), 2-O- (13%), and 3-O-subst. Rhap (7%), nonreducing end- (11%), 2-O- (10%), 3-O- (14%), and 2,6-di-O-subst. Manp units (13%). Mild reductive beta-elimination of RMP-Sp gave alpha-l-Rhap-(1-->2)-alpha-l-Rhap-(1-->3)-alpha-l-Rhap-(1-->3)-alpha-d-Manp-(1-->2)-d-Man-ol, with Man-ol substituted at O-6 with beta-d-Galp units, a related pentasaccharide lacking beta-d-Galp units, and beta-d-Galp-(1-->6)-[alpha-d-Manp-(1-->2)]-d-Man-ol in a 16:3:1w/w ratio. Traces of Man-ol and Rha-ol were detected. ESI-MS showed HexHex-ol and Hex(3-6)Hex-ol components. Three rhamnosyl units were peeled off successively from the penta- and hexasaccharide by ESI-MS-MS. The carbohydrate epitopes of RMP-Sp differ from those of the glycoprotein of Pseudallescheria boydii, a related opportunistic pathogen.International Journal of Biological Macromolecules 03/2008; 42(2):93-102. · 2.45 Impact Factor -
Article: Carbohydrates present in the glycoprotein from conidia of the opportunistic pathogen Scedosporium prolificans
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ABSTRACT: Hot aqueous extraction of conidia of Scedosporium prolificans gave a heterogeneous glycoprotein (RMP-Sp-Coni) with 41% protein and 2MeRha, Rha, Ara, Man, Gal, Glc, and GlcNH2 in a 2:18:3:47:9:15:6 M ratio, the first report of 2-O-methylrhamnose in fungi. Methylation analysis showed nonreducing end- (10%), 2-O- (11%), and 3-O-substituted Rhap (7%), nonreducing end- (8%), 2-O- (12%), 3-O- (16%), and 2,6-di-O-substituted Manp (9%), nonreducing end- (4%), 3-O- (7%), and 4-O-substituted Glcp (7%), and nonreducing end-units of Galp (9%). Mild reductive β-elimination of RMP-Sp-Coni cleaved O-linked structures to give a mixture of oligosaccharides, of which 2MeRha capping groups were present in 2MeRhaRha2Hex2Hex-ol, 2MeRhaRha2Hex-Hex-ol, 2MeRhaRha2HexHex-ol, and 2MeRhaRha2Hex-ol (ESI-MS–MS). The mixture was fractionated by Biogel P-2 column chromatography and the two predominant isolates were β-d-Galp-(1 → 6)-[2Me-α-l-Rhap-(1 → 3)-α-l-Rhap-(1 → 3)-α-d-Manp-(1 → 2)]-d-Man-ol, and another lacking the β-Galp unit. Neither was formed from mycelial glycoprotein, although β-d-Galp-(1 → 6)-[α-l-Rhap-(1 → 3)-α-l-Rhap-(1 → 3)-α-d-Manp-(1 → 2)]-d-Man-ol was a common component.Carbohydrate Polymers. -
Article: The opportunistic fungal pathogen Scedosporium prolificans: Carbohydrate epitopes of its glycoproteins
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ABSTRACT: Isolated from the mycelium of Scedosporium prolificans were complex glycoproteins (RMP-Sp), with three structurally related components (HPSEC). RMP-Sp contained 35% protein and 62% carbohydrate with Rha, Ara, Man, Gal, Glc, and GlcNH2 in a 18:1:24:8:6:5 molar ratio. Methylation analysis showed mainly nonreducing end- of Galp (13%), nonreducing end- (9%), 2-O- (13%), and 3-O-subst. Rhap (7%), nonreducing end- (11%), 2-O- (10%), 3-O- (14%), and 2,6-di-O-subst. Manp units (13%). Mild reductive β-elimination of RMP-Sp gave α-l-Rhap-(1 → 2)-α-l-Rhap-(1 → 3)-α-l-Rhap-(1 → 3)-α-d-Manp-(1 → 2)-d-Man-ol, with Man-ol substituted at O-6 with β-d-Galp units, a related pentasaccharide lacking β-d-Galp units, and β-d-Galp-(1 → 6)-[α-d-Manp-(1 → 2)]-d-Man-ol in a 16:3:1 w/w ratio. Traces of Man-ol and Rha-ol were detected. ESI-MS showed HexHex-ol and Hex3–6Hex-ol components. Three rhamnosyl units were peeled off successively from the penta- and hexasaccharide by ESI-MS-MS. The carbohydrate epitopes of RMP-Sp differ from those of the glycoprotein of Pseudallescheria boydii, a related opportunistic pathogen.International Journal of Biological Macromolecules.
